The Ste20p family of protein kinases are pleiotropic in action, influencing intracellular signaling networks through actions on multiple signal transduction pathways. We will focus our continuing studies of mammalian Ste20p-related kinases on two groups of these enzymes that we previously identified, WNKs (with no lysine (K)) and TAOs (thousand-and-one residues), which have important biological functions. The urgency of a mechanistic examination of WNKs is heightened by the discovery that they are mutated in pseudohypoaldosteronism type II, PHA II, a Mendelian form of hypertension. Lifton et al. found that mutations in either WNK1 or WNK4 causes pseudohypoaldosteronism type II, a heritable form of hypertension. Our investigation of mechanisms linking WNK1 to sodium reabsorption may reveal strategies to develop anti-hypertensive therapies. We will define the biochemical mechanisms that control WNK1 activity, and we will examine the ligands and cellular events that stimulate WNK1 activity in distal convoluted tubule cells. We have identified two targets of WNKI. We will define the biochemical mechanisms regulating the target proteins and determine functional changes associated with their phosphorylation. TAOs are the only MAP3Ks currently known to be required for activation of the p38 MAPK pathway by heterotrimeric G proteins. TAOs are highly expressed in brain, activated during myoblast differentiation, and overexpressed in some tumors, suggesting several key sites of action. We will define the biochemical mechanisms by which heterotrimeric G proteins signal to TAO2 and p38. We also found that TAO2 phosphorylates G protein alpha subunits and alters their GTP binding. Therefore, we will investigate the functional effects of phosphorylation of G proteins by TAO2, and we will determine the events in mammalian cells for which TAOs are required.